Aggrecan is a major extracellular component of articular cartilage. It is a proteoglycan responsible for providing cartilage with its mechanical properties of compressibility and elasticity. The loss of aggrecan has been implicated in the degradation of articular cartilage in arthritic diseases such as osteoarthritis.
Aggrecan contains two N-terminal globular domains, G1 and G2, separated by a proteolytically-sensitive interglobular domain, followed by a glycosaminoglycan attachment region and a C-terminal globular domain, G3. At least two enzymatic cleavage sites have been identified within the interglobular domain of aggrecan. One enzymatic cleavage site within the interglobular domain of aggrecan (asn341-phe342) has been observed to be cleaved by several known metalloproteases. Cleavage at a second aggrecan cleavage site within aggrecan (glu373-ala374) has been attributed to aggrecanase activity. The cleavage site (glu373-ala374) is therefore referred to as the aggrecanase cleavage site.
A number of aggrecanases have been cloned in recent years. These enzymes belong to a subfamily of zinc metalloproteases referred to as “ADAMTS,” an abbreviation for A Disintegrin-like And Metalloprotease domain with ThromboSpondin type I motifs. The ADAMTS family currently consists of 19 members that are related to one another on the basis of their common domain structure. Typical ADAMTS proteins contain a classic signal sequence upstream of a pro-sequence ending in a furin cleavage site, a metalloprotease domain that is well conserved among family members, a disintegrin-like motif whose functional relevance is still unknown, and at least one thrombospondin type I (TSP 1) domain. ADAMTS family members differ in the number of TSP-1 domains they contain, which can range from 1 to 15 (Cal et al., 2002; Somerville et al., 2003). The most diverse region of the ADAMTS sequence is the ‘spacer’ domain located downstream of a cysteine-rich region containing 10 structurally conserved cysteine residues. ADAMTS proteins are capable of associating with components of the extracellular matrix through interactions within the spacer domain and the TSP-1 motif(s) (Kuno and Matsushima, 1998; Tortorella et al., 2000).
ADAMTS4 (aggrecanase-1) is synthesized by IL-1 stimulated cartilage (Tortorella, et al., Science, 284:1664–1666, 1999) and is related to the degradation of aggrecan during degenerative joint diseases such as osteoarthritis (Abbaszade et al., J Biol Chem, 274: 23443–23450, 1999). ADAMTS4 is also involved in the cleavage of brain-enriched hyaluranan binding (BEHAB)/brevican, a protein that is dramatically increased in human gliomas (Matthews et al., J. Biol. Chem. 275:22695–22703, 2000). It is thus possible to ameliorate osteoarthritis and any other ADAMTS4-related diseases by inhibiting the aggrecanase activity of ADAMTS4. However, research effects on ADAMTS4 have been hampered by the instability of purified ADAMTS4 proteins.